Distinct signature of altered homeostasis in aging rod photoreceptors: implications for retinal diseases

Background

Advanced age contributes to clinical manifestations of many retinopathies and represents a major risk factor for age-related macular degeneration, a leading cause of visual impairment and blindness in the elderly. Rod photoreceptors are especially vulnerable to genetic defects and changes in microenvironment, and are among the first neurons to die in normal aging and in many retinal degenerative diseases. The molecular mechanisms underlying rod photoreceptor vulnerability and potential biomarkers of the aging process in this highly specialized cell type are unknown.

Methodology/Principal Findings

To discover aging-associated adaptations that may influence rod function, we have generated gene expression profiles of purified rod photoreceptors from mouse retina at young adult to early stages of aging (1.5, 5, and 12 month old mice). We identified 375 genes that showed differential expression in rods from 5 and 12 month old mouse retina compared to that of 1.5 month old retina. Quantitative RT-PCR experiments validated expression change for a majority of the 25 genes that were examined. Macroanalysis of differentially expressed genes using gene class testing and protein interaction networks revealed overrepresentation of cellular pathways that are potentially photoreceptor-specific (angiogenesis and lipid/retinoid metabolism), in addition to age-related pathways previously described in several tissue types (oxidative phosphorylation, stress and immune response).

Conclusions/Significance

Our study suggests a progressive shift in cellular homeostasis that may underlie aging-associated functional decline in rod photoreceptors and contribute to a more permissive state for pathological processes involved in retinal diseases.     

Cytochrome P450 1A isoenzymes in brain cells: Expression and inducibility in cultured rat brain neuronal and glial cells

Studies initiated to determine the expression of CYP1A1/1A2 isoenzymes in the primary cultures of rat brain neuronal and glial cells revealed significant activity of CYP1A-dependent 7-ethoxyresorufin-o-dealkylase (EROD) in microsomes prepared from both rat brain neuronal and glial cells. RT-PCR and immunocytochemical studies demonstrated constitutive mRNA and protein expression of CYP1A1 and 1A2 isoenzymes in cultured neuronal and glial cells. Cultured neurons exhibited relatively higher constitutive mRNA and protein expression of CYP1A1 and 1A2 isoenzymes, associated with higher activity of EROD than the glial cells. Induction studies with 3-methylchlorantherene (MC), a known CYP1A-inducer, resulted in significant concentration dependent increase in the activity of EROD in cultured rat brain cells with glial cells exhibiting a greater magnitude of induction than the neuronal cells. This difference in the increase in enzyme activity was also observed with RT-PCR and immunocytochemical studies, indicating relatively higher increase in CYP1A1 and 1A2 mRNA as well as protein expression in the cultured glial cells when compared to the neuronal cells. The greater magnitude of induction of CYP1A1 in glial cells is of significance, as these cells are components of the blood-brain barrier and it is suggested that they have a potential role in the toxication-detoxication mechanism. Our data indicating differences in the expression and sensitivity of CYP1A1 isoenzymes in cultured rat brain cells will not only help in identifying and distinguishing xenobiotic metabolizing capability of these cells but also in understanding the vulnerability of these specific cell types towards neurotoxicants.     

An expert system for the early detection of melanoma using knowledge-based image analysis

Melanoma is the most lethal skin cancer; however, nearly all patients can be saved and cured by early detection and prompt surgical treatment. It has been demonstrated that the major diagnostic and prognostic parameters of melanoma are the vertical thickness, three-dimensional (3D) size and shape, and color of the lesion. The other characteristic features of early melanoma are irregularities in the boundary of the lesion and the appearance of nonuniform pigmentation (with a variety of color). During early stages of development of the melanoma, the changes in these parameters are very difficult to assess since no good tool exists for measuring them in situ and analyzing them for malignancy. A novel optical instrument called the "Nevoscope" has been developed to obtain multiple views of the transilluminated skin lesion from several angles. These views have been used to measure the thickness and 3D size of the skin lesion without excision. A knowledge-based image analysis and interpretation system is being developed to analyze images of the skin lesion for a set of diagnostic and prognostic features: thickness, 3D size, color and margin, boundary and surface characteristics. This analysis combined with the patient's history, such as occurrence of melanoma or dysplastic nevi in the family, life style, skin type, etc., is used by the knowledge-based expert system to detect early or potentially malignant lesions. The diagnostic and prognostic knowledge bases for the early detection of melanoma are being developed with the help of expert dermatologists and published case studies.     

Adenosine 5'-diphosphate-dependent subunit dissociation of bovine dopamine beta-hydroxylase

In a previous study, it was shown that purified soluble bovine dopamine beta-hydroxylase exhibits pH-dependent reversible tetramer-dimer dissociation (Saxena, A., Hensley, P., Osborne, J. C., Jr., and Fleming, P. J. (1985) J. Biol. Chem. 260, 3386-3392). Here we report evidence for the dissociation of this enzyme by magnesium-adenosine diphosphate independent of pH in the pH range 5-7. Quantitative binding of ADP to dopamine beta-hydroxylase revealed that there are two binding sites/dimeric species of hydroxylase and that ADP is tightly bound with a KD less than 10(-8) M. Kinetic data obtained at pH 5.5, the pH inside the chromaffin granule, shows that the apparent Km values for both the substrates tyramine and ascorbate are lowered by the presence of ADP without affecting the Vmax of the enzyme. The ADP-dependent lowering of apparent Km values results from a dissociation of the enzyme to the dimeric species which has inherently lower apparent Km values for substrates.     

Effects of carbon dioxide on ethylene production and action in intact sunflower plants

A continuous flow system was used to study the interactions between carbon dioxide and ethylene in intact sunflower (Helianthus annuus L.) plants. An increase in the concentration of carbon dioxide above the ambient level (0.033%) in the atmosphere surrounding the plants increased the rate of ethylene production, and a decrease in carbon dioxide concentration resulted in a decrease in the rate of ethylene production. The change in the rate of ethylene production was evident within the first 15 minutes of the carbon dioxide treatment. Continuous treatment with carbon dioxide was required to maintain increased rate of ethylene production. The rate of carbon dioxide fixation increased in response to high carbon dioxide treatment up to 1.0%. Further increases in carbon dioxide concentration had no additional effect on carbon dioxide fixation. Carbon dioxide concentrations higher than 0.11% induced hyponasty of the leaves whereas treatment with 1 microliter per liter ethylene induced epinasty of the leaves.     

Evaluation of zinc in the regulation of serum T3 and T4 levels and hepatic functions in carbontetrachloride-intoxicated rats

Circulating tri-iodothyronine (T₃) and thyroxine (T₄) concentrations were determined after 6 wk of zinc treatment to carbontetrachloride (CCl₄) intoxicated male albino rats. Concentrations of T₃ were observed to be significantly depressed following CCl₄ treatment alone. On the contrary, no significant change was noticed in the concentrations of T₄ when compared to controls. However, zinc administration to hepatotoxic animals resulted in restoring the T₃ activity to within normal limits, thus indicating the indirect effects of zinc on the regulation of thyroid hormone concentrations. The activities of all the serum and hepatic marker enzymes were found to be significantly elevated following CCl₄ treatment. However, following zinc supplementation to these intoxicated animals, the levels of the marker enzymes decreased significantly when compared to the CCl₄-treated animals. A similar trend was seen in the case of lipid peroxidation following zinc treatment.